National Center for Magnetic Resonance in Wuhan, Key Laboratory of Magnetic Resonance in Biological Systems, State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics, Wuhan Institute of Physics and Mathematics, Innovation Academy for Precision Measurement Science and Technology, Chinese Academy of Sciences, Wuhan, 430071, People's Republic of China.
Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, 20892-0520, USA.
Commun Biol. 2021 Mar 5;4(1):282. doi: 10.1038/s42003-021-01802-1.
Diacylglycerol kinase (DgkA) is a small integral membrane protein, responsible for the ATP-dependent phosphorylation of diacylglycerol to phosphatidic acid. Its structures reported in previous studies, determined in detergent micelles by solution NMR and in monoolein cubic phase by X-ray crystallography, differ significantly. These differences point to the need to validate these detergent-based structures in phospholipid bilayers. Here, we present a well-defined homo-trimeric structure of DgkA in phospholipid bilayers determined by magic angle spinning solid-state NMR (ssNMR) spectroscopy, using an approach combining intra-, inter-molecular paramagnetic relaxation enhancement (PRE)-derived distance restraints and CS-Rosetta calculations. The DgkA structure determined in lipid bilayers is different from the solution NMR structure. In addition, although ssNMR structure of DgkA shows a global folding similar to that determined by X-ray, these two structures differ in monomeric symmetry and dynamics. A comparative analysis of DgkA structures determined in three different detergent/lipid environments provides a meaningful demonstration of the influence of membrane mimetic environments on the structure and dynamics of membrane proteins.
二酰基甘油激酶（DgkA）是一种小型的膜内在蛋白，负责在 ATP 依赖性条件下将二酰基甘油磷酸化为磷酸脂。此前报道的其结构分别通过溶液 NMR 在胶束中以及 X 射线晶体学在单油酸甘油酯立方相中确定，差异非常显著。这些差异表明需要在磷脂双层中验证基于去污剂的这些结构。在这里，我们通过魔角旋转固态 NMR（ssNMR）光谱，使用一种结合了分子内和分子间顺磁弛豫增强（PRE）衍生的距离约束和 CS-Rosetta 计算的方法，展示了 DgkA 在磷脂双层中的明确的同三聚体结构。在脂质双层中确定的 DgkA 结构与溶液 NMR 结构不同。此外，尽管 DgkA 的 ssNMR 结构显示出与 X 射线确定的结构相似的整体折叠，但这两种结构在单体对称性和动力学方面存在差异。对在三种不同的去污剂/脂质环境中确定的 DgkA 结构的比较分析提供了一个有意义的例证，说明了膜模拟环境对膜蛋白结构和动力学的影响。
